Mass regeneration of Coffea arabica L. somatic embryos using a temporary immersion bioreactor was improved by optimizing the immersion cycles, i.e. both the duration and the frequency of immersions. It was demonstrated that increasing the frequency of short immersions (1 min immersions every 24, 12 and 4 h) stimulated embryo production (480, 2,094 and 3,081 embryos/1-l bioreactor, respectively) and improved quality (60, 79 and 85% of torpedo shaped embryos, respectively). On the other hand, an increase in the immersion duration (1, 5 and 15 min) inhibited embryo regeneration (from 2,094 to 428 embryos per 1-l bioreactor) and negatively affected their morphological quality (from 79 to 49% torpedo-shaped embryos) and the conversion of embryos into plants (from 70 to 33%). A 15 min immersion duration applied every 4 h produced hyperhydric symptoms in 90% of the embryos. Hyperhydric embryos were characterized by higher fresh weight and water content, more negative values for water potential and higher K + content when compared to normal torpedo-shaped embryos. Micrographs showed structural problems in the globular stage, such as the existence of an irregular epidermis and an absence of reserves. Whatever the immersion cycle used, the somatic embryos exhibited water and mineral characteristics very different from those of their zygotic counterparts. The use of 1 min immersions every 4 h led to the production of the largest quantities of torpedo-shaped embryos without hyperhydricity that succeeded in regenerating plants (75% conversion).
<strong>Título en ingles: Micropropagation of papaya plants in temporary immersion recipients from axilary shoots</strong><p><strong>Título corto: Micropropagación de lechosa en recipientes de inmersión temporal.</strong></p><strong>Resumen: </strong>Se estandarizaron las condiciones de iniciación, multiplicación, enraizamiento y aclimatización de plantas hermafroditas de lechosa cv Maradol provenientes de brotes axilares, producidos en recipientes de inmersión temporal RITA<sup>®</sup>. En cada envase, contentivo de 200 ml de medio de cultivo líquido de Fitch, se colocaron cuatro brotes de 2 a 3 cm de longitud. Los biorreactores se conectaron a tres líneas de inmersión de 5, 2 y 1 min cada 4h y se colocaron 6 envases en promedio por línea, en condiciones de fotoperíodo de 16 h. Transcurridos 30 a 45 días, se cuantificaron los brotes y se clasificaron de acuerdo al tamaño: < 2 cm (pequeños), entre 2 a 3 cm (medianos), ˃ 3 cm con y sin raíz (grandes). Los dos primeros tipos de brotes se continuaron multiplicando en los mismos medios; y los más elongados se aclimatizaron utilizando el Sistema Autotrófico Hidropónico (SAH). Se determinó la sanidad y la fidelidad de las plantas producidas mediante pruebas de ELISA y RAPD, respectivamente. Durante un periodo de 6 meses se reciclaron un total de 47 recipientes, los cuales produjeron 1.091 brotes: 377 pequeños; 482 medianos; 175 grandes sin raíz y 57 con raíz. Usando el SAH se obtuvo 89,5% de plantas aclimatizadas cuando se usaron brotes enraizados, y 41,6% a partir de brotes sin raíces. Con la combinación de las técnicas RITA y SAH se logró un sistema continuo y eficiente de producción de plantas sanas y fieles al tipo, en comparación con los métodos convencionales de micropropagación y aclimatización.<p><strong>Palabras clave</strong>: <em>Carica papaya</em>, RITA<sup>®</sup>, sistema autotrófico, estabilidad genética.</p><p><strong>Abstract: </strong>We standardized initiation, multiplication, rooting and acclimatization conditions of papaya cv Maradol hermaphrodite plants from axillary buds produced in temporary immersion reactor RITA<sup>®</sup>. Recipients contained 200 ml of Fitch liquid culture medium, and four shoots of 2 to 3 cm. in length were placed in each. The bioreactors were connected to three different immersion lines of 5, 2, and 1 min each 4h, with 6 containers per line on average, in 16 h photoperiod. After 30 to 45 days, the shoots produced were quantified and classified according to size: <2 cm (small), from 2 to 3 cm (medium), >3 cm with or without roots (large). The first two types of shoots were multiplied in the same culture media, and more elongated shoots were acclimatized using Autotrophic Hydroponic System (AHS). The sanity and fidelity of the produced plants were determined using ELISA and RAPD, respectively. For a period of six months 47 vessels were recycled and 1,091 shoots were produced: 377 small; 482 medium; 175 large without roots and 57 rooted shoots. Using AHS, 89.5% acclimatized plants were obtained when rooted shoots were used, and 41.6% from rootless shoots. With the combination of RITA and AHS techniques we achieved a continuous and efficient production of healthy and true to type papaya plants, in comparison to conventional micropropagation and acclimatization procedures.</p><p><strong>Key words</strong>: <em>Carica</em><em> papaya</em>, RITA<sup>®</sup>, autotrophic system, genetic stability.</p><p><strong>Recibido:</strong> mayo 16 de 2014<strong> Aprobado: </strong>abril 21 de 2015</p>
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